Project Summary This application addresses NCI PQ6 on how circadian processes affect tumor development by studying the role of circadian dysfunction of nuclear receptor pathways in non-alcoholic fatty liver disease (NAFLD)- induced hepatocellular carcinoma (HCC). HCC, previously considered a rare cancer in the Western world, has increased 3-fold in incidence since the 1980s, and is currently the fastest rising cause of cancer-related death in the U.S, due to an increase in incidence in combination of lack of understanding of its mechanism. The increased HCC risk is coupled with the prevalence of obesity-related NAFLD, which has recently become the leading risk factor for HCC. However, no efficient approaches for prevention, early diagnosis, and treatment of NAFLD-induced HCCs are currently available. The increased risk of NAFLD-related HCC is coupled with population-wide chronic circadian disruption, a common risk factor of obesity, NAFLD, and cancer in humans. We have recently established a jet-lagged mouse model following a human nightshift schedule. We found that chronic jet-lag induces metabolic syndrome, NAFLD, and HCC in wild-type mice following a pathophysiological pathway strikingly similar to that observed in obese humans, with NAFLD progressing to nonalcoholic steatohepatitis (NASH) and fibrosis prior to HCC detection. We identified intrahepatic cholestasis as the key pathophysiological mechanism promoting NAFLD-induced HCC, and demonstrated that circadian dysregulation of nuclear receptor FXR and CAR pathways is an essential oncogenic mechanism that drives cholestasis and toxic bile acid signaling to promote the progression from NAFLD to NASH, fibrosis, and eventually HCC. In this application, we propose 3 aims. 1) Define the circadian profiles of FXR and CAR controlled hepatic gene networks and jet-lag induced deregulation signatures of these gene networks. 2) Define the role of Ctnnb1 and Tp53 and other oncogenic mutations in the progression from jet-lag induced liver premalignant lesions to HCC and the circadian profiles of gene signatures as well as serum biomarkers associated with hepatocarcinogenesis. 3) Test the predicted abilities of FXR agonist obeticholic acid (OCA) and CAR inverse agonist androstanol (ANDR) to prevent jet-lag induced cholestasis and HCC. These studies will significantly improve our understanding of the role of circadian dysfunction in spontaneous carcinogenesis in general, and NFALD-induced hepatocarcinogenesis in particular. They will also illuminate new and exciting avenues to develop novel chronotherapy strategies for prevention and treatment of NAFLD-induced HCC in humans.